The objective of this 2-year R21 application is to extend the capabilities of the Flp-FRTsite specific recombinase system (pioneered in mice by my laboratory) by generating inducible tools that will permit the generation of temporally controlled genetic modifications in embryonic and adult mice; notably, tools that can be used in parallel with either constitutive or inducible Cre-loxP-based reagents. This resulting dual recombinase capability is not presently available and would represent an important advance, permitting the engineering of either successive gene modifications as a means to model multistep developmental or adult disease processes or simultaneous gene modifications as a means to model multigene disorders. This advanced capability to temporally regulate recombination will be especially important for study of processes occurring at late embryonic and adult stages, those processes most limited by standard (non-inducible) conditional approaches. The proposed experiments focus on generating two types of inducible Flp tools, one broadly active and the other tissue-specific. These experiments follow from our previous results showing that the Flp variant, Flpe, is as effective in mice as Cre, and from the work of others showing that steroid-receptor fusions with Cre enable temporal regulation of recombination. In aim 1, we will introduce into mice widely inducible Flpe transgenes capable of catalyzing genetic changes in cells of diverse tissues at chosen times in embryonic and adult life. In aim 2, we will introduce into mice a tissue-specific inducible Flpe transgene capable of mediating genetic modifications in both a spatially and temporally regulated fashion in the neural tube. These mice will permit the genetic modification of specific subsets of cell lineages, distinguished by their temporal emergence from the dorsal neural tube. Moreover, they will serve as a prototype for other tissue-specific inducible Flpe transgenics. While the proposed tools will provide new capabilities for delineating processes critical to the development and health of many organ systems, our particular interest centers on brainstem lineages, in particular, those lineages that emerge successively from the rhombic lip region of the dorsal hindbrain. We plan to apply the proposed tools in conjunction with Cre-loxP-containing reagents to manipulate "late-born" rhombic lip lineages critical to the formation of both the precerebellar afferent system, a set of diverse nuclei essential for motor control, and the medullary raphe system, a center for homeostatic control in which developmental abnormalities have been implicated in sudden infant death syndrome.